Rapid detection and differentiation of Taenia species are required for the control and prevention of taeniasis and cysticercosis in areas where these diseases are endemic. Because of the lower ...sensitivity and specificity of the conventional diagnosis based on microscopical examination, molecular tools are more reliable for differential diagnosis of these diseases. In this study, we developed and evaluated a loop-mediated isothermal amplification (LAMP) assay for differential diagnosis of infections with Taenia species with cathepsin L-like cysteine peptidase (clp) and cytochrome c oxidase subunit 1 (cox1) genes. LAMP with primer sets to the cox1 gene could differentiate between three species, and LAMP with primer sets to the clp gene could differentiate Taenia solium from Taenia saginata/Taenia asiatica. Restriction enzyme digestion of the LAMP products from primer set Tsag-clp allowed the differentiation of Taenia saginata from Taenia asiatica. We demonstrated the high specificity of LAMP by testing known parasite DNA samples extracted from proglottids (n = 100) and cysticerci (n = 68). LAMP could detect one copy of the target gene or five eggs of T. asiatica and T. saginata per gram of feces, showing sensitivity similar to that of PCR methods. Furthermore, LAMP could detect parasite DNA in all taeniid egg-positive fecal samples (n = 6). Due to the rapid, simple, specific, and sensitive detection of Taenia species, the LAMP assays are valuable tools which might be easily applicable for the control and prevention of taeniasis and cysticercosis in countries where these diseases are endemic.
During 2012-2013, a total of 4325 host-seeking adult ticks belonging to the genus Ixodes were collected from various localities of Hokkaido, the northernmost island of Japan. Tick lysates were ...subjected to real-time PCR assay to detect borrelial infection. The assay was designed for specific detection of the Relapsing fever spirochete Borrelia miyamotoi and for unspecific detection of Lyme disease-related spirochetes. Overall prevalence of B. miyamotoi was 2% (71/3532) in Ixodes persulcatus, 4.3% (5/117) in Ixodes pavlovskyi and 0.1% (1/676) in Ixodes ovatus. The prevalence in I. persulcatus and I. pavlovskyi ticks were significantly higher than in I. ovatus. Co-infections with Lyme disease-related spirochetes were found in all of the tick species. During this investigation, we obtained 6 isolates of B. miyamotoi from I. persulcatus and I. pavlovskyi by culture in BSK-M medium. Phylogenetic trees of B. miyamotoi inferred from each of 3 housekeeping genes (glpQ, 16S rDNA, and flaB) demonstrated that the Hokkaido isolates were clustered with Russian B. miyamotoi, but were distinguishable from North American and European B. miyamotoi. A multilocus sequence analysis using 8 genes (clpA, clpX, nifS, pepX, pyrG, recG, rplB, and uvrA) suggested that all Japanese B. miyamotoi isolates, including past isolates, were genetically clonal, although these were isolated from different tick and vertebrate sources. From these results, B. miyamotoi-infected ticks are widely distributed throughout Hokkaido. Female I. persulcatus are responsible for most human tick-bites, thereby I. persulcatus is likely the most important vector of indigenous relapsing fever from tick bites in Hokkaido.
Trematodes of the family Dicrocoeliidae commonly use terrestrial mollusks as the first intermediate host. Despite abundant studies on the adult worms in birds and mammals, few reports exist on their ...larval stage in snail intermediate hosts. A present survey of mollusks in Japan led us to the discovery of dicrocoeliid sporocysts with cercariae in 16 of 303 individuals, encompassing 8 snail species and 1 slug species. A DNA barcoding based on sequencing of mitochondrial cytochrome c oxidase subunit 1 showed that the larvae consisted of 5 species. Phylogenetic trees of nuclear 18S and 28S ribosomal DNAs confirmed the 5 species to be members of the Dicrocoeliidae. These were temporarily termed dicrocoeliid species 1 to 5, because conclusive identification was impossible without adult worms. These unknown species were phylogenetically related to each other, except for sp. 5. The phylogenetic trees demonstrated close genetic relationships between sp. 3 and the genus Lutztrema and between sp. 5 and the genus Lyperosomum. The phylogenetic analysis also suggests a possibility that the currently accepted macrotaxonomy of the Dicrocoeliidae is problematic, due to the paraphyly of the subfamilies Dicrocoeliinae and Leipertrematinae. Morphological characterization of the cercariae and their DNA barcodes provide a primary platform for differentiating dicrocoeliids from various mollusks in Japan. The DNA barcodes, in particular, will enable tracing the parasite life cycles, in case of finding metacercariae and adults from presently unknown hosts.
Abstract Fasciolosis is an economically important disease of livestock caused by Fasciola hepatica , Fasciola gigantica , and aspermic Fasciola flukes. The aspermic Fasciola flukes have been ...discriminated morphologically from the two other species by the absence of sperm in their seminal vesicles. To date, the molecular discrimination of F. hepatica and F. gigantica has relied on the nucleotide sequences of the internal transcribed spacer 1 (ITS1) region. However, ITS1 genotypes of aspermic Fasciola flukes cannot be clearly differentiated from those of F. hepatica and F. gigantica. Therefore, more precise and robust methods are required to discriminate Fasciola spp. In this study, we developed PCR restriction fragment length polymorphism and multiplex PCR methods to discriminate F. hepatica , F. gigantica , and aspermic Fasciola flukes on the basis of the nuclear protein-coding genes, phosphoenolpyruvate carboxykinase and DNA polymerase delta, which are single locus genes in most eukaryotes. All aspermic Fasciola flukes used in this study had mixed fragment pattern of F. hepatica and F. gigantica for both of these genes, suggesting that the flukes are descended through hybridization between the two species. These molecular methods will facilitate the identification of F. hepatica , F. gigantica , and aspermic Fasciola flukes, and will also prove useful in etiological studies of fasciolosis.
Taenia saginata
is the most common species of tapeworm infecting humans. Infection is acquired by eating cysticercus larvae in undercooked beef. A closely related species,
T. asiatica
, is found in ...eastern and southeastern Asia. The larvae of
T. asiatica
develop in viscera of pigs. In northern Russia, there is a third member of this morphologically indistinguishable group. Cysticerci of so-called northern
T. saginata
are found in cerebral meninges of reindeer, and the unique life cycle is dependent on a native custom of eating raw reindeer brain. We report the winding history of this mysterious tapeworm from the first reports to the present time. In addition, we confirm the position of this parasite as a strain of
T. saginata
by analyzing a mitochondrial DNA sequence of an archival specimen. The origin of this strain might date back to reindeer domestication and contacts between cattle-herding and reindeer-herding peoples in Asia.
Understanding the ecology of ticks and tick-borne microorganisms is important to assess the risk of emerging tick-borne diseases. Despite the fact that the
tick bites humans, we lack information ...including population genetics and the reason for the inadequate distribution in Japan. A 5-year survey revealed that Rishiri Island, the main stopover in the East Asian Flyway of wild birds in the northern Sea of Japan, was a refuge of
. The
included two haplogroups, which were supposed to diverge a long time before the island separated from the continent and Hokkaido mainland. The detection of microorganisms from wildlife revealed that wild birds and rodents play a role in diffusion and settlement, respectively, of not only
but also
-borne microorganisms including
Ehrlichia khabarensis and
US lineage. Various island-specific factors control
dominance and tick-borne pathogen maintenance. The results may enable us to explain how tick-borne infectious microorganisms are transported.
Purpose
We aimed to evaluate the magnetic resonance imaging (MRI) appearance changes during stereotactic radiotherapy (SRT) for large sized brain metastases, and analyze the lesions necessitating ...treatment plan modification.
Materials and methods
A total of 23 patients (27 lesions, >2 cm in tumor diameter) underwent SRT and all lesions were evaluated the appearance changes which had the necessity of the treatment plan modification. The appearance change of tumor during SRT was evaluated using gadolinium-enhanced MRI. The reasons of the modification were classified into tumor reduction, tumor enlargement, displacement, and shape change.
Results
Among the 27 lesions, 55.6% required the treatment plan modification. The reasons were tumor reduction in six lesions, tumor enlargement in three lesions, displacement in three lesions, and shape change in three lesions. The planning target volume (PTV) size changed up to 43.0% and the shift of center of PTV was a maximum of 1.7 mm. The pathological status (adenocarcinoma vs others) and timing of steroid administration (prior vs after SRT start) were the predictive factors of tumor changes required the modification.
Conclusions
As tumor changes might occur even during short period of SRT, the treatment plan evaluation and modification were important in SRT for large brain metastases.
Abstract
Daily dose distributions for adaptive radiotherapy (ART) using helical tomotherapy (HT) are calculated using megavoltage computed tomography (MVCT). Generally, the MVCT number is converted ...to mass density (MD) using an MD calibration table (MVCT-MD table). The aims of this study are to calculate the tolerance levels of the MD for ART and to evaluate the tolerance levels using clinical patient plans. These tolerance levels of MD were calculated based on the tissue maximum ratio (TMR) of 6MV flattening-filter-free (FFF) beam of HT and the effective tissue thickness data from an International Commission on Radiological Protection 110 phantom data for lung, adipose/muscle and cartilage/spongy-bone. These tolerance levels were determined by considering both the MD causing a dose error of 2% and the variation in MVCT numbers. Subsequently, the stability of the MD values was estimated with the standard deviations (SD) in the MVCT number over 6 months. The dose distribution for clinical patient plans was calculated using the MVCT-MD table with added tolerance levels. These tolerance levels were determined as MD differences causing a dose error of 2%, and were ± 0.049 g/cm3, ± 0.030 g/cm3 and ± 0.049 g/cm3 for lung, adipose/muscle and cartilage/spongy-bone, respectively. The calculated dose distribution errors using the MVCT-MD table added tolerance levels were within 2%. We proposed these tolerance levels in MD for the quality control of the MVCT-MD table.
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